scholarly journals Monitoring Nuclear Transport in HeLa Cells Using the Green Fluorescent Protein

BioTechniques ◽  
1996 ◽  
Vol 21 (1) ◽  
pp. 62-63 ◽  
Author(s):  
Shivani Chatterjee ◽  
Ursula Stochaj
Keyword(s):  
Green Fluorescent Protein ◽  
Hela Cells ◽  
Fluorescent Protein ◽  
Nuclear Transport ◽  
Green Fluorescent

A novel yellowish-green fluorescent protein from the marine copepod, Chiridius poppei, and its use as a reporter protein in HeLa cells

Gene ◽  
2006 ◽  
Vol 372 ◽  
pp. 18-25 ◽  
Author(s):  
Hiromi Masuda ◽  
Yasuhiro Takenaka ◽  
Atsushi Yamaguchi ◽  
Satoshi Nishikawa ◽  
Hiroshi Mizuno
Keyword(s):  
Green Fluorescent Protein ◽  
Hela Cells ◽  
Fluorescent Protein ◽  
Reporter Protein ◽  
Marine Copepod ◽  
Yellowish Green ◽  
Green Fluorescent

scholarly journals Cortactin Is Necessary for F-Actin Accumulation in Pedestal Structures Induced by Enteropathogenic Escherichia coli Infection

2002 ◽  
Vol 70 (4) ◽  
pp. 2206-2209 ◽  
Author(s):  
Vlademir V. Cantarelli ◽  
Akira Takahashi ◽  
Itaru Yanagihara ◽  
Yukihiro Akeda ◽  
Kinichi Imura ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Green Fluorescent Protein ◽  
Hela Cells ◽  
Fluorescent Protein ◽  
Enteropathogenic Escherichia Coli ◽  
Escherichia Coli Infection ◽  
Green Fluorescent ◽  
Pedestal Formation

ABSTRACT Cortactin and the translocated intimin receptor, Tir, interacted with each other in pedestal formation in HeLa cells infected with enteropathogenic Escherichia coli (EPEC). Cortactin is shown to be necessary for organizing actin pedestals in response to EPEC, based on the expression of green fluorescent protein-fused cortactin derivatives in HeLa cells.

scholarly journals Functional Regions of the Pseudomonas aeruginosa Cytotoxin ExoU

2005 ◽  
Vol 73 (1) ◽  
pp. 573-582 ◽  
Author(s):  
Shira D. P. Rabin ◽  
Alan R. Hauser
Keyword(s):  
Amino Acids ◽  
Pseudomonas Aeruginosa ◽  
Green Fluorescent Protein ◽  
Amino Acid ◽  
Hela Cells ◽  
Fluorescent Protein ◽  
Host Cells ◽  
Phospholipase Activity ◽  
C Terminus ◽  
Green Fluorescent

ABSTRACT ExoU, a potent patatin-like phospholipase, causes rapid cell death following its injection into host cells by the Pseudomonas aeruginosa type III secretion system. To better define regions of ExoU required for cytotoxicity, transposon-based linker insertion mutagenesis followed by site-directed mutagenesis of individual residues was employed by using a Saccharomyces cerevisiae model system. Random insertion of five amino acids identified multiple regions within ExoU that are required for cell killing. Five regions were chosen for further characterization: three corresponded to the oxyanion hole, hydrolase motif, and catalytic aspartate motif of the patatin-like domain within the N-terminal half of ExoU; one corresponded to an uncharacterized part of the patatin-like domain; and one corresponded to a region near the C terminus. Specific individual amino acid substitutions in each of the four N-terminal regions prevented killing of yeast and significantly reduced phospholipase activity. Whereas five amino acid insertions in the fifth region near the C terminus markedly reduced cytotoxicity and phospholipase activity, substitution of individual amino acids did not abolish either activity. To determine whether each of the five identified regions of ExoU was also essential for cytotoxicity in human cells, representative mutant forms of ExoU fused to green fluorescent protein were expressed in HeLa cells. These variants of ExoU were readily visualized and caused minimal cytotoxicity to HeLa cells, while wild-type ExoU fused to green fluorescent protein induced significant cell lysis and no detectable fluorescence. Thus, a minimum of five regions, including one which is well removed from the patatin-like domain, are required for the cytotoxicity and phospholipase activity of ExoU.

scholarly journals Fluorescence lifetime images of green fluorescent protein in HeLa cells during TNF-α induced apoptosis

2009 ◽  
Vol 8 (6) ◽  
pp. 763 ◽  
Author(s):  
Toshiyuki Ito ◽  
Shugo Oshita ◽  
Takakazu Nakabayashi ◽  
Fan Sun ◽  
Masataka Kinjo ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescence Lifetime ◽  
Hela Cells ◽  
Fluorescent Protein ◽  
Tnf Α ◽  
Induced Apoptosis ◽  
Green Fluorescent

scholarly journals A functional chimaeric S-layer-enhanced green fluorescent protein to follow the uptake of S-layer-coated liposomes into eukaryotic cells

2004 ◽  
Vol 379 (2) ◽  
pp. 441-448 ◽  
Author(s):  
Nicola ILK ◽  
Seta KÜPCÜ ◽  
Gerald MONCAYO ◽  
Sigrid KLIMT ◽  
Rupert C. ECKER ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Fusion Protein ◽  
Hela Cells ◽  
Enhanced Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Laser Scanning Microscopy ◽  
Host Cells ◽  
Confocal Laser ◽  
Green Fluorescent ◽  
Egfp Fluorescence

The chimaeric gene encoding a C-terminally truncated form of the S-layer protein SbpA of Bacillus sphaericus CCM 2177 and the EGFP (enhanced green fluorescent protein) was ligated into plasmid pET28a and cloned and expressed in Escherichia coli. Just 1 h after induction of expression an intense EGFP fluorescence was detected in the cytoplasm of the host cells. Expression at 28 °C instead of 37 °C resulted in clearly increased fluorescence intensity, indicating that the folding process of the EGFP moiety was temperature sensitive. To maintain the EGFP fluorescence, isolation of the fusion protein from the host cells had to be performed in the presence of reducing agents. SDS/PAGE analysis, immunoblotting and N-terminal sequencing of the isolated and purified fusion protein confirmed the presence of both the S-layer protein and the EGFP moiety. The fusion protein had maintained the ability to self-assemble in suspension and to recrystallize on peptidoglycan-containing sacculi or on positively charged liposomes, as well as to fluoresce. Comparison of fluorescence excitation and emission spectra of recombinant EGFP and rSbpA31-1068/EGFP revealed identical maxima at 488 and 507 nm respectively. The uptake of liposomes coated with a fluorescent monomolecular protein lattice of rSbpA31-1068/EGFP into HeLa cells was studied by confocal laser-scanning microscopy. The major part of the liposomes was internalized within 2 h of incubation and entered the HeLa cells by endocytosis.

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